PTH is a primary regulator of calcium homeostasis and bone metabolism, and its signaling system has served as a major target for the development of novel anabolic therapeutic approaches for osteoporosis. However, the exact mechanisms by which PTH exerts its actions in bone are not fully understood. Despite the progress having been made in determining the PTH downstream signals in osteoblasts, novel signaling components may exist to bridge some major gaps in our understanding of the bone anabolic effects of PTH. We have characterized a novel PTH signaling pathway in osteoblasts. Our preliminary data show that binding of PTH to its receptor PTH1R induced association of LRP6, a coreceptor of Wnt, with PTH1R. The formation of the ternary complex containing PTH, PTH1R and LRP6 promoted rapid LRP6 phosphorylation, which resulted in the recruitment of axin to LRP6, and stabilization of 2-catenin. Activation of PKA is essential for PTH-induced 2-catenin stabilization, but not for Wnt signaling. Our demonstration of that the LRP6 coreceptor mediates PTH- activated 2-catenin signaling in osteoblasts suggests for the first time that LRP6 is the component of PTH/PTH1R complex. Importantly, in vivo studies confirmed that PTH treatment led to phosphorylation of LRP6 and an increase in amount of 2-catenin in osteoblasts with a concurrent increase in bone formation in rat. Therefore, we hypothesize that PTH-induced phosphorylation of LRP6, which is induced through activation of PKA, is required for PTH anabolic effect on bone. The objective of this proposal is to characterize the mechanisms of the LRP6 activation in response to PTH stimulation in osteoblasts and examine the role of LRP6 in PTH bone anabolic action in vivo. The proposal is organized into three aims. In aim I, the PKA phosphorylation sites within LRP6 will be characterized, and the role of PKA-phosphorylated LRP6 in activation of LRP6-2-catenin signaling in osteoblasts will be determined. In aim II, the requirement of LRP6 in mediating PTH-stimulated osteoblast activities in primary calvarial preosteoblasts and bone marrow stromal cells will be defined. In aim III, the requirement of LRP6 in PTH bone anabolic action in vivo will be examined using a mouse model of OC- Cre-mediated conditional deletion of LRP6. PUBLIC HEALTH RELEVANCE: The mechanisms responsible for anabolic actions of PTH on bone are not completely understood. This proposal will characterize how LRP6, a coreceptor for Wnt, is activated in osteoblasts upon PTH stimulation and the role of LRP6 in PTH bone anabolic action.